A Structural And Functional Basis For The Regulation Of Gene Expression By Nuclear Retention Of RNA
Funder
National Health and Medical Research Council
Funding Amount
$504,097.00
Summary
The nuclear retention mechanism is a novel way used by cells to control which genes are made into proteins - a fundamental process for all diseases, particularly cancers. This project will employ cutting edge structural and proteomic techniques to determine the molecular details underpinning nuclear retention. These insights will be important for the development of new tissue-restricted gene therapy applications and drugs targeting the cancers that rely on this mechanism.
Post Transcriptional Regulation Of The Plasminogen Activator Inhibitor Type 2 Gene
Funder
National Health and Medical Research Council
Funding Amount
$241,527.00
Summary
The process of wound healing, removal of blood clots, cell migration and the metastatic spread of cancers requires the recruitment of specialised proteases. These proteases act primarily to degrade other proteins, mainly in the extracellular space, which in turn allow cells to move around, wounds to close, and blood clots to disappear. The plasminogen activating system is one of the most important enzyme systems involved in these events. One of the proteases that cleaves plasminogen to its activ ....The process of wound healing, removal of blood clots, cell migration and the metastatic spread of cancers requires the recruitment of specialised proteases. These proteases act primarily to degrade other proteins, mainly in the extracellular space, which in turn allow cells to move around, wounds to close, and blood clots to disappear. The plasminogen activating system is one of the most important enzyme systems involved in these events. One of the proteases that cleaves plasminogen to its active form, plasmin, is urokinase (u-PA). Plasminogen activator inhibitor type 2 (PAI-2) is a serine protease inhibitor that inhibits u-PA activity. The degree of u-PA activity therefore depends on the relative levels of u-PA and PAI-2. In addition to controlling u-PA activity, PAI-2 also influences intracellular events including cell proliferation, differentiation and apoptosis. PAI-2 protein and mRNA levels are substantially modulated by many cytokines and growth factors. This project addresses the molecular mechanisms underlying the regulation of PAI-2 gene expression. We have recently shown that a significant degree of PAI-2 regulation occurs at the level of PAI-2 mRNA stability, and we have identified two regions within the PAI-2 mRNA that play a role in this process. Both regions provide binding sites for cellular proteins. We have identified one of these binding proteins to be HuR, a protein that has recently been shown to control the stability of other mRNAs. The specific aims of this project are firstly, to determine the role of HuR in the control of PAI-2 mRNA stability, and secondly, to clone a characterise the other PAI-2 mRNA binding proteins we have identifed. An understanding of how cells modulate levels of PAI-2 mRNA will significantly add to the broader field of gene regulation and may also provide new clues to influence PAI-2 levels in the body.Read moreRead less
A New Window into Transgene Silencing in Plants: mechanisms of copy-number independent, 5' sequence dependent, post-transcriptional silencing in a complex polyploid. Silencing of introduced genes is a major problem limiting plant molecular improvement. Sugarcane, a complex polyploid, shows the most efficient transgene silencing ever observed in plants. Silencing operates on the RNA, depends on the upstream sequence of the gene, and is independent of copy number. Other plant species develop endop ....A New Window into Transgene Silencing in Plants: mechanisms of copy-number independent, 5' sequence dependent, post-transcriptional silencing in a complex polyploid. Silencing of introduced genes is a major problem limiting plant molecular improvement. Sugarcane, a complex polyploid, shows the most efficient transgene silencing ever observed in plants. Silencing operates on the RNA, depends on the upstream sequence of the gene, and is independent of copy number. Other plant species develop endopolyploidy with age, and show unpredictable or patchy silencing. We speculate that differential silencing is a natural control mechanism in the exploitation of polyploidy in plants. The sugarcane system provides an exceptional opportunity to identify the sequences that trigger and protect from silencing, and to develop approaches to avoid the problem.Read moreRead less
I am a molecular biologist determining the mechanisms of eukaryotic mRNA translation and its regulation by RNA-binding proteins and noncoding RNA. In collaborative work I extend these basic science objectives into the medical research areas of cardiology
Post-transcriptional Regulation Of Plasminogen Activator Inhibitor 2 Gene Expression
Funder
National Health and Medical Research Council
Funding Amount
$508,838.00
Summary
Plasminogen activator inhibitor type 2 (PAI-2) is a protease inhibitor that has intracellular and extracellular functions. The PAI-2 gene is highly regulated at the level of PAI-2 mRNA stability. We have identified regions within the PAI-2 transcript essential for this regulation and a number of novel proteins that engage these regions. This project is aimed at understanding how these and other proteins control PAI-2 expression at the mRNA level.
Expression and characterisation of nutrient transporters from the intracellular malaria parasite, Plasmodium falciparum. The malaria parasite invades the red blood cells of its host and this provides it with a safe haven in which to grow and replicate. Within the red blood cell, the parasite takes up nutrients and excretes metabolic wastes via specialised membrane transport proteins which are, as yet, very poorly understood. The sequencing of the malaria parasite genome has enabled us to ident ....Expression and characterisation of nutrient transporters from the intracellular malaria parasite, Plasmodium falciparum. The malaria parasite invades the red blood cells of its host and this provides it with a safe haven in which to grow and replicate. Within the red blood cell, the parasite takes up nutrients and excretes metabolic wastes via specialised membrane transport proteins which are, as yet, very poorly understood. The sequencing of the malaria parasite genome has enabled us to identify candidates for a wide variety of these proteins. The aim of this project is to establish systems in which the functional properties of these transporter proteins may be characterised in detail.Read moreRead less
Molecular Mechanisms For The Cell-type Specific Regulation Of The Tissue-type Plasminogen Activator Gene
Funder
National Health and Medical Research Council
Funding Amount
$490,500.00
Summary
Tissue-type plasminogen activator (t-PA) is an important enzyme that is widely known for its ability to remove blood clots. More recently, t-PA has been shown to influence memory development and under pathological conditions can promote neuronal cell death. t-PA is produced by many cells including the endothelial cells that line the blood vessels, fibroblasts, as well as cells within the central nervous system. The t-PA gene is regulated very differently in these cell types and this project will ....Tissue-type plasminogen activator (t-PA) is an important enzyme that is widely known for its ability to remove blood clots. More recently, t-PA has been shown to influence memory development and under pathological conditions can promote neuronal cell death. t-PA is produced by many cells including the endothelial cells that line the blood vessels, fibroblasts, as well as cells within the central nervous system. The t-PA gene is regulated very differently in these cell types and this project will address the mechanisms underlying the cell-type specific regulation of the t-PA gene. Endothelial cells, fibroblasts and neuronal cell cultures will be used to study the regulation of t-PA expression. Information gained will not only add to the understanding of the broader field of gene regulation, but may also provide clues to manipulate the expression of the t-PA gene in different cells.Read moreRead less
Inherited disorders of the blood, such as sickle-cell anaemia and thalassaemia, result from mutations in the genes that produce haemoglobin. Current treatments can partially alleviate some of the debilitating symptoms of these diseases but these treatments have significant side effects, and despite the best efforts of clinicians, many patients succumb to their conditions at an early age. It has been observed that certain individuals exhibit a milder form of the disease, as a consequence of the r ....Inherited disorders of the blood, such as sickle-cell anaemia and thalassaemia, result from mutations in the genes that produce haemoglobin. Current treatments can partially alleviate some of the debilitating symptoms of these diseases but these treatments have significant side effects, and despite the best efforts of clinicians, many patients succumb to their conditions at an early age. It has been observed that certain individuals exhibit a milder form of the disease, as a consequence of the reactivation of their foetal haemoglobin genes, (a distinct set of genes that would have been active in utero but are normally silenced around the time of birth). It is widely accepted that if pharmaceutical means can be found for reactivating the foetal haemoglobin genes then many patients would benefit. The regulation of the foetal globin genes, like most human genes, is complicated and there are few obvious means of increasing their activity. Nevertheless, it is believed that by investigating the molecular mechanisms by which they are controlled it will be possible to devise therapeutic agents that mimic these mechanisms or to develop agents that prevent the shutdown of the foetal genes around birth. To this end we have been working on the molecules that regulate the activity of the haemoglobin genes. We have recently cloned a number of DNA-binding proteins, and their co-factors, that appear to be involved in silencing foetal globin gene expression. This grant proposal is concerned with learning how these new molecules operate to silence gene expression as a first step towards designing agents that will prevent the silencing.Read moreRead less
Plant transformation: exploiting anti-apoptosis genes for very high efficiency transformation. Crop improvement through genetic modification depends on the ability to transform target species. The most desirable method is Agrobacterium mediated transformation. However, plant species and cultivars differ significantly in their ability to be efficiently transformed by Agrobacterium. This is particularly true for the economically important cereals. We have discovered that anti-apoptosis genes, whic ....Plant transformation: exploiting anti-apoptosis genes for very high efficiency transformation. Crop improvement through genetic modification depends on the ability to transform target species. The most desirable method is Agrobacterium mediated transformation. However, plant species and cultivars differ significantly in their ability to be efficiently transformed by Agrobacterium. This is particularly true for the economically important cereals. We have discovered that anti-apoptosis genes, which inhibit programmed cell death, dramatically increase the Agrobacterium transformation efficiency in bananas and sugarcane. We will utilise this information and develop the use of these genes to increase the efficiency of transformation in those crops and cultivars that are difficult to transform using Agrobacterium.Read moreRead less
Translational Control Of Gene Expression And The Choice Between Cell Death And Proliferation
Funder
National Health and Medical Research Council
Funding Amount
$378,000.00
Summary
Proteins carry out most enzymatic and structural functions in a cell. Thus, the kinds of protein molecules that are found in a given cell determine its characteristics and cells respond to changes in their environment by adjusting the abundance of some or many proteins in their collection. The instructions for the assembly of proteins are encoded in the genes and this information is expressed via intermediary molecules called messenger (m)RNA. Both, transcription of the genes into mRNA molecules ....Proteins carry out most enzymatic and structural functions in a cell. Thus, the kinds of protein molecules that are found in a given cell determine its characteristics and cells respond to changes in their environment by adjusting the abundance of some or many proteins in their collection. The instructions for the assembly of proteins are encoded in the genes and this information is expressed via intermediary molecules called messenger (m)RNA. Both, transcription of the genes into mRNA molecules and their subsequent translation by the ribosomes into protein are tightly controlled steps in the gene expression pathway. Erroneous gene expression is a major factor in human disease and dysregulation of translation is linked to a growing spectrum of illnesses such as cancer and cardiovascular disease, viral infection, and less frequent hereditary syndromes. The project proposed here is prompted by emerging evidence for a role of translational regulation in controlling the balance between cell death and survival. Tipping this balance has disastrous consequences for an organism as evidenced by its involvement in many major disorders (e. g. stroke, heart failure, neurodegeneration, AIDS, cancer, autoimmunity). Our aim is to test the hypothesis that a putative translational regulator termed p97-DAP5-NAT1, and a specialised mechanism of translation initiation by internal ribosome entry are important for the maintenance of this balance. To investigate this, we will employ DNA chips, a novel tool from Genomics research that allows the measurement of the levels of thousands of mRNA molecules in a single experiment. It is conceivable that knowledge of these special mechanisms of translation will lead to novel targets for therapeutic intervention, and this work will contribute some of the experimental tools to explore these avenues in the future.Read moreRead less